Oyster shell hydrogen powder composition with pH buffering function and preparation method thereof

Abstract

The present invention discloses an oyster shell hydrogen powder composition with pH buffering function. The composition is composed of the following components by mass percentage: 40% to 50% oyster shell hydrogen powder, 20% to 25% starch, 10% to 15% citric acid, and 15% to 20% hydroxyethyl methylcellulose, with the sum of the percentages of the above components being 100%. The present invention also discloses a method for preparing the above oyster shell hydrogen powder composition. The composition and preparation method disclosed in the present invention solve the problem that existing products containing oyster shell hydrogen powder, when releasing H2, simultaneously produce Ca(OH)2, which is strongly alkaline and can cause significant irritation and damage to the gastrointestinal mucosa.

Description

Technical Field

[0001] The invention belongs to the technical field of medicines and health care products, and particularly relates to an oyster shell hydrogen powder composition with a pH buffering function. The invention also relates to a preparation method of the composition. Background Art

[0002] Oyster shell powder (OSP), also known as shell powder, is made from dried and ground mussel, oyster, clam, and snail shells. Its main ingredients are calcium carbonate, calcium gluconate (96.4%), and pure calcium (38.0%). It also contains trace elements such as magnesium, iron, manganese, and phosphorus. Its main benefits include: ① Calcium supplementation: OSP's high calcium content promotes bone development, improves bone toughness and density, and effectively prevents osteoporosis. ② Analgesia: OSP is a traditional Chinese medicine with astringent and analgesic properties. It effectively inhibits free acid secretion, reduces acid irritation in the gastric mucosa, prevents gastroenteritis, and accelerates the healing of ulcers. ③ Sedation: The active ingredients in OSP act directly on the central nervous system to produce a calming and tranquilizing effect, effectively alleviating symptoms such as insomnia and neurasthenia.

[0003] H2 is the smallest molecule in nature. It is colorless, odorless, and harmless. It possesses strong penetrating power, rapid diffusion, and the ability to penetrate various biological barriers, while maintaining exceptional biosafety. In recent years, H2 has garnered increasing attention as a molecule with a wide range of biological activities. Over 1,000 domestic and international research papers on hydrogen medicine reveal its numerous health benefits. The most fundamental of these is its selective antioxidant properties. It helps slow the aging process of the skin, providing cosmetic and anti-aging benefits. It helps regulate metabolism and alleviate metabolic diseases. It helps activate the immune system to boost immunity. It helps inhibit cell mutation and reduce apoptosis, promoting the repair and regeneration of damaged tissues. It helps reduce inflammatory responses, stabilize cell membranes, alleviate allergic symptoms, and reduce radiation damage. It can also help relieve fatigue and improve work efficiency and quality of life. The 2014 "National Food Safety Standard for the Use of Food Additives" (GB2760-2014) lists H2 as a food additive. It can enter the body through inhalation or oral administration, freely binding to cells and exerting its antioxidant, anti-inflammatory, and anti-apoptotic properties. With the development of hydrogen molecular medicine, H2 has garnered widespread attention as a medical gas in therapeutic research for a variety of diseases. The human body produces a limited amount of H2, normally producing approximately 150ml per day. This is primarily generated by intestinal hydrogen-producing bacteria fermenting unabsorbed carbohydrates, making it difficult to meet long-term antioxidant needs. Despite the promising medical applications of H2, existing methods for preparing products such as hydrogen-rich water and beverages primarily generate H2 through water electrolysis or the reaction of metallic magnesium with water. These methods often suffer from issues such as unstable dissolved H2 levels and the tendency for H2 to escape.

[0004] Oyster shell hydrogen powder (OSHP) is made by enriching oyster shell powder with a specific amount of hydrogen (H2) through a special physical process. The main component of this H2-enriched powder is calcium hydride (CaH2), also known as hydrogenated OSP. While retaining OSP's calcium-strengthening, bone-strengthening, astringent, and sedative properties, OSHP releases a significant amount of H2 upon contact with water after oral administration, enhancing H2's various health benefits, including antioxidant, anti-inflammatory, immune-boosting, blood circulation-boosting, sleep-enhancing, and wound-healing properties. Studies have demonstrated that OSHP continuously releases H2 when contacted with water, both in vitro and in vivo. Both the amount of H2 released and the duration of this release exceed those of commercially available hydrogen-rich water (H2-rich aqueous solutions). Unfortunately, OSHP reacts chemically with water (H2O) according to the chemical equation: CaH2+2H2O=2H2↑+Ca(OH)2. The Ca(OH)2 produced while releasing H2 is a strong alkaline substance, which can cause significant irritation and damage to the gastrointestinal mucosa. The safety of OSHP limits its oral development and application. Summary of the Invention

[0005] The first object of the present invention is to provide an oyster shell hydrogen powder composition with pH buffering function. This solves the problem that existing products containing oyster shell hydrogen powder release H2 while producing Ca(OH)2, which is strongly alkaline and can cause significant irritation and damage to the gastrointestinal mucosa.

[0006] The second object of the present invention is to provide a method for preparing the above composition.

[0007] The first technical solution adopted by the present invention is: an oyster shell hydrogen powder composition with a pH buffering function is composed of the following components by mass percentage: 40% to 50% oyster shell hydrogen powder, 20% to 30% starch, 10% to 15% citric acid, and 15% to 20% hydroxyethyl methylcellulose, the sum of the percentages of the above components being 100%.

[0008] The first technical solution adopted by the present invention is also characterized in that:

[0009] Furthermore, the oyster shell hydrogen powder composition with the function of buffering pH is composed of the following components by mass percentage: 45% oyster shell hydrogen powder, 24% starch, 13% citric acid, and 18% hydroxyethyl methylcellulose.

[0010] Furthermore, citric acid acts as an acidity regulator to adjust the pH value of the composition to a stable neutral state.

[0011] Furthermore, hydroxyethyl methylcellulose is used as a sustained-release preparation to improve the stability of the composition and enhance the safety of the composition.

[0012] Furthermore, the oyster shell hydrogen powder composition is packaged in capsule shells, the capsule shells are made of edible and easily digestible gelatin, and the capsules are made into capsules with a content of 0.45 g per capsule.

[0013] The second technical solution adopted by the present invention is: a method for preparing an oyster shell hydrogen powder composition with a pH buffering function, which is prepared according to the following steps:

[0014] Step 1: Pass oyster shell hydrogen powder, starch, citric acid, and hydroxyethyl methylcellulose through a 100-mesh sieve respectively for later use;

[0015] Step 2: According to the formula, accurately weigh the oyster shell hydrogen powder, starch, citric acid, and hydroxyethyl methylcellulose and mix them together;

[0016] Step 3, filling the total mixed material into hollow capsules;

[0017] Step 4: Pack the filled capsules.

[0018] The beneficial effects of the present invention are:

[0019] 1. The oyster shell hydrogen powder composition of the present invention has significant anti-inflammatory and wound healing effects, and can provide the body with hydrogen in a long-term and stable manner. Oral administration of an OSHP prescription combination with a buffered pH value can effectively increase the H2 content in the human body, thereby playing an anti-inflammatory and wound healing role. In addition, an OSHP prescription combination with a buffered pH value also has the advantages of high safety and minimal side effects, making it suitable for long-term use by a wide range of people.

[0020] 2. The present invention utilizes solid-state hydrogenation technology to produce a hydrogen-rich OSHP, which slowly and continuously releases hydrogen upon contact with water. This allows for the slow and continuous release of hydrogen on the body surface after external application, promoting rapid wound healing.

[0021] 3. The present invention utilizes solid-state hydrogenation technology to produce an H2-rich oyster shell powder, which slowly and continuously releases H2 upon contact with water. Oral administration through the gastrointestinal tract can slowly and continuously release H2 with certain health benefits, thereby achieving significant anti-inflammatory, immune and metabolic regulation effects and promoting rapid wound healing.

[0022] 4. The composition of the present invention uses OSHP, citric acid, hydroxyethyl methylcellulose (HEMC), and starch as ingredients in an OSHP formulation with a buffered pH. Leveraging OSHP's ability to release hydrogen over a long period of time, citric acid (CA) is added for acid-base buffering, resulting in an OSHP formulation with an anti-inflammatory, immune-modulating, metabolic-regulating, and wound-healing promoting pH adjusted to 7. Hydroxyethyl methylcellulose (HEMC) is further added to improve drug stability and enhance sustained-release properties, thereby enhancing drug safety.

[0023] 5. The present invention's pH-buffered oyster shell hydrogen powder composition immediately neutralizes and buffers the strongly alkaline Ca(OH)2 produced after oral OSHP ingestion, generating calcium citrate (Ca(C6H5O6)2), an organic calcium that is more easily absorbed by the body. While releasing hydrogen, the liquid pH is consistently maintained at around 7, preventing Ca(OH)2 from irritating or damaging the gastrointestinal mucosa. This invention proposes a pH-buffered OSHP formulation. This technical feature scientifically combines the biological health benefits of hydrogen with calcium supplementation, resulting in a synergistic health effect. The technology is highly mature, and pilot trials have confirmed that it meets the requirements for large-scale industrial production.

[0024] 6. The powdering process of the composition preparation method of the present invention adopts a mechanical powdering method, using a high-speed grinder to grind the raw materials such as oyster hydrogen powder and citric acid into powder. This method is faster and more efficient than traditional methods, while also ensuring a uniform and fine powder. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is a trend diagram of H2 release concentration over 26 hours for compositions of Example 1 of the present invention with different OSHP contents;

[0026] Figure 2 This is a 26-hour pH value trend diagram of the composition of Example 1 of the present invention when different contents of OSHP release H2;

[0027] Figure 3A This is a schematic diagram showing comparative observation of the anti-inflammatory and anti-allergic effects of OSHP and dexamethasone in the composition of the present invention on mice with atopic dermatitis;

[0028] Figure 3B is a schematic diagram comparing the effects of OSHP and dexamethasone in the composition of the present invention on the scores of atopic dermatitis mice;

[0029] Figure 3C This is a schematic diagram comparing the effects of OSHP and dexamethasone in the composition of the present invention on the scratching frequency of mice with atopic dermatitis;

[0030] Figure 3Dis a schematic diagram comparing the effects of OSHP and dexamethasone in the composition of the present invention on body weight changes in mice with atopic dermatitis;

[0031] Figure 3E This is a schematic diagram comparing the effects of OSHP and dexamethasone in the composition of the present invention on changes in spleen coefficients in mice with atopic dermatitis;

[0032] Figure 4A This is a schematic diagram comparing the effects of OSHP and dexamethasone in the composition of the present invention on the pathological morphology of skin ulcers in mice with atopic dermatitis;

[0033] Figure 4B This is a schematic diagram comparing the effects of OSHP and dexamethasone in the composition of the present invention on the degree of skin ulcers in mice with atopic dermatitis;

[0034] Figure 4C 1 is a schematic diagram showing the comparative results of the effects of OSHP and dexamethasone in the composition of the present invention on the pathological morphology of epidermal hyperplasia in mice with atopic dermatitis;

[0035] Figure 4D 2 is a schematic diagram showing the comparison of the effects of OSHP and dexamethasone in the composition of the present invention on the degree of epidermal hyperplasia in mice with atopic dermatitis;

[0036] Figure 5A 3 is a schematic diagram showing the comparison of the effects of OSHP and dexamethasone in the composition of the present invention on serum IgE levels in mice with atopic dermatitis;

[0037] Figure 5B 1 is a schematic diagram showing the comparison of the effects of OSHP and dexamethasone in the composition of the present invention on the serum IL-17 level in mice with atopic dermatitis;

[0038] Figure 6A 1 is a schematic diagram comparing the effects of OSHP and dexamethasone in the composition of the present invention on the absolute number of lymphocytes in the peripheral blood of mice with atopic dermatitis;

[0039] Figure 6B 1 is a comparative diagram of the effects of OSHP and dexamethasone in the composition of the present invention on the absolute number of monocytes in the peripheral blood of mice with atopic dermatitis;

[0040] Figure 6C 1 is a comparative diagram of the effects of OSHP and dexamethasone in the composition of the present invention on the absolute number of neutrophils in the peripheral blood of mice with atopic dermatitis;

[0041] Figure 6D is a comparative schematic diagram of the effects of OSHP and dexamethasone in the composition of the present invention on the proportion of lymphocytes in the peripheral blood of mice with atopic dermatitis;

[0042] Figure 6E FIG1 is a comparative diagram of the effects of OSHP and dexamethasone in the composition of the present invention on the proportion of monocytes in the peripheral blood of mice with atopic dermatitis;

[0043] Figure 6F is a comparative schematic diagram of the effects of OSHP and dexamethasone in the composition of the present invention on the proportion of neutrophils in the peripheral blood of mice with atopic dermatitis;

[0044] Figure 7 Schematic diagram of the observation results of the back wound recovery effects of scalded mice in each group of the present invention;

[0045] Figure 8 This is a schematic diagram comparing the weight changes of scalded mice in each group of the present invention. DETAILED DESCRIPTION

[0046] The present invention will be further described below with reference to the accompanying drawings and specific embodiments.

[0047] The present invention provides an oyster shell hydrogen powder composition with a buffered pH value. The composition, comprising an OSHP buffering agent, can instantly adjust the strong alkaline substance Ca(OH)2 in the process of CaH2+2H2O=2H2↑+Ca(OH)2 through an acid-base buffer, and a preparation method thereof. The pH value can be sustainably stabilized at approximately 7, resulting in a neutral pH. The preparation method of the present invention is simple and easy to implement, and is suitable for mass industrial production.

[0048] The present invention addresses the bottleneck problem of the existing product OSHP, which produces Ca(OH)2 when releasing H2, which strongly alkalines the gastrointestinal mucosa and causes obvious irritation and damage. It provides a scientific solution and preparation method that have been verified by multiple experiments, and relates to the field of food and health products. The raw materials are: oyster shell hydrogen powder, starch, citric acid, and hydroxyethyl methylcellulose. Experiments have shown that the present invention can continuously release H2 for 26 hours after mixing OSHP with water, and the pH value of the liquid is stable at around 7. Through animal experimental studies, the results revealed that the OSHP prescription combination significantly improves the symptoms of atopic dermatitis and has a good final therapeutic effect; the OSHP prescription combination accelerates the healing of wounds on scalded mice, repairs the affected skin, and promotes the growth and healing of wounds. The present invention adopts the following technical solution: according to the specification of 0.45g of content per capsule and 1000 capsules, the components are composed of the following components by mass percentage: 40% to 50% of oyster shell hydrogen powder, 20% to 25% of starch, 10% to 15% of citric acid, and 15% to 20% of hydroxyethyl methylcellulose, and the sum of the percentages of the above components is 100%.

[0049] The present invention also provides a method for preparing an oyster shell hydrogen powder composition with a buffered pH value, comprising the following steps:

[0050] (1) Pass OSHP, starch, citric acid, and hydroxyethyl methylcellulose through a 100-mesh sieve respectively;

[0051] (2) According to the formula, OSHP, starch, citric acid, and hydroxyethyl methylcellulose were accurately weighed and mixed;

[0052] (3) filling the total mixed material of (2) into hollow capsules;

[0053] (4) Packing the filled capsules.

[0054] The OSHP formulation of the present invention is a hydrogen-ion-rich oyster shell extract powder produced domestically using solid-state hydrogenation technology. Upon contact with water, it releases large amounts of hydrogen over a long period of time. pH regulator selection: To control the pH of the formulation and facilitate hydrogen release, a pH regulator is required. Initially, sodium dihydrogen phosphate was used, but this resulted in uneven mixing. Furthermore, considering that the addition of large amounts of phosphorus could affect calcium absorption and thus the effectiveness of the formulation, citric acid was ultimately selected. Citric acid is more acidic and can more effectively adjust the pH value. The resulting calcium citrate is also more easily absorbed in the body. This improves drug utilization without causing stomach damage. The innovation of this invention lies in the acidity regulator, citric acid, with the molecular formula C6H8O6, a weak organic acid. When the Ca(OH)2 in OSHP meets the citric acid aqueous solution, calcium citrate (Ca(C6H5O6)2) reacts with water (H2O) to form calcium citrate, a chemical reaction: Ca(OH)2 + 2C6H8O6 = Ca(C6H5O6)2 + 2H2O. This neutralization reaction is unidirectional and has no secondary reactions. Calcium citrate is also an organic calcium acid with excellent absorption and is non-irritating to the gastrointestinal mucosa, further enhancing its calcium supplementation, analgesic, and anti-inflammatory effects. Hydroxyethyl methylcellulose (HEMC) is a nonionic surfactant with auxiliary and sustained-release properties, including thickening, suspending, binding, floating, dispersing, and film-forming properties. It improves drug stability and solubility, controlling the drug release rate and achieving a sustained-release effect. Starch, as a fundamental excipient in oral solid dosage forms, acts as a filler to increase volume and a disintegrant to promote rapid drug release. Starch's adhesive, dispersing, and lubricating properties are crucial for ensuring the stability and homogeneity of formulations. Its hygroscopicity provides additional protection for drugs, effectively resisting moisture damage and maintaining their stability. Starch also improves the drug's taste. The innovation of the powdering process lies in the use of a mechanical powdering method, using a high-speed grinder to grind raw materials such as oyster hydrogen powder and citric acid. This method is faster and more efficient than traditional methods, while also ensuring a uniform and fine powder.

[0055] The technical solution of the present invention is further described below through examples and drawings.

[0056] Example 1

[0057] This embodiment adopts the following technical solution: according to the specification of 0.45g content / capsule, 1000 capsules are made, and the components and amounts are as follows: 30 parts (45%) of oyster shell hydrogen powder, 16 parts (24%) of starch, 8.8 parts (13%) of citric acid, and 12 parts (18%) of hydroxyethyl methylcellulose.

[0058] The preparation method comprises the following steps:

[0059] (1) Pass OSHP, starch, citric acid, and hydroxyethyl methylcellulose through a 100-mesh sieve respectively;

[0060] (2) According to the formula, OSHP, starch, citric acid, and hydroxyethyl methylcellulose were accurately weighed and mixed;

[0061] (3) filling the total mixed material of (2) into hollow capsules;

[0062] (4) Packing the filled capsules.

[0063] Example 2

[0064] This embodiment adopts the following technical scheme: according to the specification of 0.45g content / capsule, 1000 capsules are made, and the components and amounts are as follows: 26.7 parts (40%) of oyster shell hydrogen powder, 20 parts (30%) of starch, 10 parts (15%) of citric acid, and 10 parts (15%) of hydroxyethyl methylcellulose.

[0065] The preparation method comprises the following steps:

[0066] (1) Pass OSHP, starch, citric acid, and hydroxyethyl methylcellulose through a 100-mesh sieve respectively;

[0067] (2) According to the formula, OSHP, starch, citric acid, and hydroxyethyl methylcellulose were accurately weighed and mixed;

[0068] (3) filling the total mixed material of (2) into hollow capsules;

[0069] (4) Packing the filled capsules.

[0070] Example 3

[0071] This embodiment adopts the following technical solution: according to the specification of 0.45g content / capsule, 1000 capsules are made, and the components and amounts are as follows: 33.4 parts (50%) of oyster shell hydrogen powder, 13.4 parts (20%) of starch, 6.7 parts (10%) of citric acid, and 13.4 parts (20%) of hydroxyethyl methylcellulose.

[0072] The preparation method comprises the following steps:

[0073] (1) Pass OSHP, starch, citric acid, and hydroxyethyl methylcellulose through a 100-mesh sieve respectively;

[0074] (2) According to the formula, OSHP, starch, citric acid, and hydroxyethyl methylcellulose were accurately weighed and mixed;

[0075] (3) filling the total mixed material of (2) into hollow capsules;

[0076] (4) Packing the filled capsules.

[0077] The effects of the composition of the present invention are verified by experiments in conjunction with the accompanying drawings. The following embodiments are for illustrative purposes and are not intended to limit the scope of the present invention. Other embodiments based on the embodiments of the present invention that are not innovatively developed are intended to fall within the scope of protection of the present invention.

[0078] The present invention provides an OSHP formulation with a buffered pH value, comprising: a capsule shell made of edible and easily digestible gelatin, and capsules with a content of 0.45 g per capsule. The capsule contents contain the following ingredients: oyster shell hydrogen powder, starch, citric acid, and hydroxyethyl methylcellulose.

[0079] like Figure 1 As shown, the hydrogen-enriched effect of the composition provided in Example 1 was verified:

[0080] During the experiment, the same distilled water, the same temperature, the same measuring instrument, and the same technical standard measurement by the same person were used to ensure that there was only one variable, the H2 release amount of the powder containing the hydrogen ion-rich OSHP prescription combination. Figure 1 A line graph shows the hydrogen release in water from a hydrogen-rich OSHP formulation at different concentrations: 1 mg / mL, 5 mg / mL, 10 mg / mL, 20 mg / mL, and 100 mg / mL. The graph shows that the OSHP formulation can continuously release hydrogen for 26 hours. During the first 6 hours, H2 release slowly increases across the different concentrations of the OSHP formulation with a buffered pH. After 6 hours, hydrogen release gradually decreases, reaching zero at the 26th hour. The graph shows that H2 release is related to the concentration of the capsule powder. Higher concentrations increase hydrogen release, reaching a maximum of 523 ppb in the 100 mg / mL group. Within 16 hours of the experiment, H2 release in all five concentration groups consistently exceeded 200 ppb.

[0081] like Figure 2 As shown, the acid-base buffering effect of the composition provided in Example 1 was verified:

[0082] During the experiment, the same distilled water, the same temperature, the same measuring instrument, and the same person were used to perform the same technical standard measurements to ensure that there was only one variable, pH, for the hydrogen ion-rich OSHP formulation combination. Figure 2 The figure is a line graph of the pH values ​​of the powder of a hydrogen ion-rich OSHP prescription combination in water at different concentrations, namely 1 mg / mL, 5 mg / mL, 10 mg / mL, 20 mg / mL and 100 mg / mL. From the figure, it can be seen that the pH value of a hydrogen ion-rich OSHP prescription combination can be continuously stabilized at 7 during the process of continuously releasing H2 in water.

[0083] Experiment 1: Experimental study on the anti-inflammatory effect of the composition of Example 1 on atopic dermatitis

[0084] Atopic dermatitis (AD) is a chronic skin disease also known as atopic eczema or hereditary allergic dermatitis. Clinically, it manifests as an eczematous rash on the localized skin, accompanied by significant itching and exudation. Scratching may also result in bleeding and erosion. Exudation gradually forms scabs and scales, severely impacting the quality of life of patients and their families. Currently, the treatment of AD focuses on avoiding allergens, relieving clinical symptoms, and reducing complications and recurrence rates. Mild AD patients generally use topical corticosteroids (TCS) and calcineurin inhibitors (TCL), while moderate and severe AD patients, in addition to maintaining TCS and TCL, also require antihistamines, cyclosporine, and phototherapy to control pruritus. Experiment 1 established an AD mouse model by inducing 2,4-dinitrochlorobenzene (DNCB) to evaluate the improvement of the OSHP prescription combination on AD mice, providing a theoretical basis for verifying the anti-inflammatory and anti-allergic effects of the OSHP prescription combination.

[0085] 1. Reagent Preparation

[0086] Preparation of 0.5% sodium carboxymethyl cellulose: Dissolve 1g of sodium carboxymethyl cellulose in 150ml of hot water and place in a hot magnetic stirrer for high temperature dissolution. After all the solids are dissolved, the volume is adjusted to 200ml.

[0087] Acetone and olive oil mixed solution: Acetone and olive oil are evenly mixed in a ratio of 3:1 by volume.

[0088] 1% DNCB solution: Weigh 1.2 g of DNCB solid and dissolve it in 12 ml of a mixed solution of acetone and olive oil (9 ml of acetone + 3 ml of olive oil), and mix thoroughly.

[0089] 0.4% DNCB solution: Weigh 48 mg of DNCB solid and dissolve it in 12 ml of a mixture of acetone and olive oil (9 ml of acetone + 3 ml of olive oil), and mix thoroughly.

[0090] OSHP prescription combination solution: effective concentration 50 mg / kg / d, dissolved in 0.5% sodium carboxymethyl cellulose.

[0091] 2. DNCB animal specific dermatitis modeling, grouping and drug administration

[0092] The experiment was divided into four groups: normal group, model group, dexamethasone group, and OSHP combination group. A 2.5 cm x 2.5 cm area of ​​mouse back skin was shaved with an electric shaver. Mice with problematic back skin were removed to avoid interference. The shaved area was then depilated with a depilatory cream to ensure complete exposure of the mouse's back skin. After using the depilatory cream, the mice were washed with water to prevent corrosion of the depilatory cream on the mouse skin and affect the modeling effect. On day 1 of the experiment, the model group, dexamethasone positive control group, and OSHP combination group were sensitized with 1% DNCB solution (200 μL per mouse). The normal group was sensitized with a mixture of acetone and olive oil (3:1 acetone:olive oil) for 3 consecutive days. On day 8, atopic dermatitis was induced by applying 200 μL of 0.4% DNCB solution to the back of the mice. The normal control group was sensitized with a mixture of acetone and olive oil (3:1 acetone:olive oil) for 4 consecutive days. Drug treatment was started on the day of modeling. All groups were given drugs by gavage for 12 consecutive days, and experimental samples were collected on the 13th day.

[0093] 3. Changes in atopic dermatitis scores

[0094] According to the AD scoring standard, the severity of skin lesions on the back of mice was graded, and the severity of lesions was scored according to the presence of erythema / hemorrhage, scratches / erosions, skin edema, scaling, or lichenification on the back. Each level of severity was scored from 0 to 4 points. The scores of different levels were added together to obtain the final score. Scores were scored every two days according to the above method. Higher scores indicate more severe symptoms. The average value was finally taken. The scoring content and standards are as follows:

[0095] (1) Skin erythema / hemorrhage: 4 points if the area is greater than 75% of the exposed skin area; 3 points if the area is between 40% and 75%; 2 points if the area is between 10% and 40%; 1 point if the area is less than 10%; 0 points if there is no bleeding or erythema.

[0096] (2) Skin scratches / erosions: 4 points if the area is larger than 75% of the exposed skin area; 3 points if the area is between 40% and 75%; 2 points if the area is between 10% and 40%; 1 point if the area is less than 10%; 0 points if there is no bleeding or erythema.

[0097] (3) Skin edema: dark red skin, moist surface, obvious swelling, and no new hair were scored as 4 points; light red skin, slightly dry surface, reduced edema, and no new hair were scored as 3 points; pink skin, slightly darker than normal mice, with an uneven surface and new hair, scored as 2 points; skin color close to the normal skin color of mice, with a large amount of new hair and smooth skin were scored as 1 point; none of the above symptoms were scored as 0 points.

[0098] (4) Scaly or lichenified skin: 4 points if the area is larger than 75% of the exposed skin area; 3 points if the area is between 40% and 75%; 2 points if the area is between 10% and 40%; 1 point if the area is less than 10%; 0 points if there is no bleeding or erythema.

[0099] like Figure 3A 、 Figure 3B As shown, the skin of mice in the normal group was smooth and free of dermatitis, with a dermatitis score of 0. The dermatitis score in the model group was relatively high (P<0.05). Erythema and edema appeared on the back skin. Compared with the model group, the dermatitis scores of mice in the dexamethasone and OSHP combination groups were lower. The erythema on the back of the mice was reduced, the desquamation was improved, and the area of ​​scabs gradually decreased. The OSHP combination and dexamethasone groups showed a good therapeutic effect on atopic dermatitis in mice. The back skin of the mice did not show redness or swelling, but only small patches of erythema and scales. Compared with the model group, the symptoms of mice in the treatment groups were significantly improved, demonstrating that the OSHP combination has a good therapeutic effect on atopic dermatitis.

[0100] 4. Changes in behavioral index scores (number of scratching) of AD mice

[0101] The scratching behavior of AD mice is also an important indicator of the severity of mouse dermatitis. The observation method is to place them alone in a transparent mouse cage ten minutes after DNCB sensitization and observe the scratching behavior of the mice on the back and ears. A series of one or more scratching movements of the mouse's hind paws toward the ears is counted as one scratch. The scratch ends when the mouse's hind paws are placed back on the floor. The time is counted for 10 minutes, the data is recorded, and the average value is finally taken.

[0102] like Figure 3C As shown in the figure, the mice in the normal group had smooth skin and no scratching behavior because they were not sensitized. The model group showed obvious scratching behavior with a very high frequency, indicating that the mice had a high degree of skin inflammation. The dexamethasone group and the OSHP capsule group scratched less frequently than the model group and showed significant improvement.

[0103] 5. Changes in spleen index in AD mice

[0104] Mice were killed by dislocation and the spleen was placed on filter paper for weighing. The spleen index was calculated as spleen mass (mg) / mouse body weight (mg). The larger the spleen index, the more severe the inflammatory response of the mouse. Therefore, the spleen index is also an indicator of the severity of mouse atopic dermatitis.

[0105] like Figure 3D 、 Figure 3E As shown, the spleen index of mice in the OSHP combination group and the dexamethasone group was lower than that in the model group, indicating that the OSHP combination can reduce inflammation in mice and play a role in treating atopic dermatitis.

[0106] 6. Pathological changes of skin tissue in AD area of ​​mice

[0107] On the 13th day of the experiment, the mice were killed by dislocation, and the damaged skin on the back was removed and fixed with 4% paraformaldehyde. The skin was routinely dehydrated, embedded in paraffin, sectioned, and stained with hematoxylin-eosin (HE). The pathological changes of the back skin tissue were observed under an optical microscope.

[0108] like Figure 4A 、 Figure 4B 、 Figure 4C 、 Figure 4D As shown, the dorsal skin tissue of mice in the normal group was normal and showed no significant changes. The skin structure of all layers was intact, the dermis and epidermis were clearly demarcated, and no inflammatory cell infiltration was observed. However, the dorsal skin of mice in the model group showed severe hyperkeratosis, acanthosis, and a large number of inflammatory cells infiltrating the upper dermis, which fully conforms to the pathogenesis of AD. Compared with the model group, the pathological tissue results of mice in the OSHP combination group showed reduced epidermal keratinization and less severe dermatitis lesions.

[0109] 7. Effects of OSHP prescription combination on cytokines in AD skin lesions

[0110] At the end of the experiment, when collecting mouse tissues, we collected blood from the mouse eyeballs, extracted the mouse serum after static centrifugation, and placed it in a -80 degree refrigerator. Subsequently, we used ELISA to detect the blood sample concentrations of Ig E and IL-17 in the mouse serum. By comparing the concentrations of Ig E and IL-17 between different groups, we compared the degree of inflammation in the mice, and reflected the therapeutic effect of the OSHP prescription combination on atopic dermatitis by the concentration of immune factors in the serum. This experiment mainly used Mouse Immunoglobulin E (IgE) ELISA Kit and Mouse IL-17 ELISA Kit to detect the expression levels of Ig E and IL-17, respectively.

[0111] like Figure 5A 、 Figure 5B As shown in the data, compared with the model group, the expression levels of Ig E and IL-17 in the OSHP prescription combination group and the positive control dexamethasone group were significantly lower than those in the model group after treatment. Therefore, the OSHP prescription combination may reduce the release of inflammatory factors during the occurrence of atopic dermatitis through the role of immune regulation, thereby achieving the effect of alleviating inflammation and relieving dermatitis. Compared with the positive control, it can be seen that the OSHP prescription combination and dexamethasone have basically the same effect in relieving inflammation. Therefore, the OSHP prescription combination can treat AD by reducing the expression of inflammatory factors such as Ig E and IL-17 in the serum during the inflammation process.

[0112] like Figure 6A As shown in Figure 2, the absolute number of lymphocytes in the peripheral blood of the model group mice was significantly higher than that of the normal mice. Figure 6D Compared with the model group, the absolute number of lymphocytes in the peripheral blood of the dexamethasone treatment group increased significantly, as shown in Figure 2. Figure 6A As shown in Figure 3, the OSHP prescription combination group tended to suppress the proportion and absolute number of lymphocytes in peripheral blood compared with the model group, as shown in Figure 3. Figure 6D It is noteworthy that when the OSHP prescription combination treatment group was compared with the dexamethasone treatment group, there were statistically significant differences in the proportion and absolute number of suppressed lymphocytes, as shown in Figure 2. Figure 6A and 6D Secondly, compared with the normal group mice, the proportion and absolute number of monocytes and neutrophils in the peripheral blood of the model group mice showed an upward trend, as shown in Figure 2. Figure 6B 、 Figure 6C 、 Figure 6E and Figure 6F As shown, the dexamethasone-treated group showed an inhibitory trend in the absolute number and proportion of monocytes and granulocytes (the average change was more pronounced, but due to large intra-group variability, no statistically significant difference was found). The OSHP combination group had no effect on the proportion and absolute number of monocytes and granulocytes in mice. Since lymphocytes are the main cell type in peripheral blood, accounting for 70-90% of peripheral blood mononuclear cells, it can be seen that the OSHP combination group inhibits AD mainly by suppressing the proportion and absolute number of lymphocytes and the cytokines they secrete.

[0113] The above experimental results show that the OSHP prescription combination has a significant improvement effect on the treatment of AD at a concentration of 50 mg / kg. The research results show that the OSHP prescription combination has an improvement effect on AD, can reduce the epidermal hyperplasia of AD mice, and play a role in repairing the skin of mice. In addition, the H2 released by an oyster shell hydrogen powder prescription combination with a buffered pH value in the mouse body can also play an anti-inflammatory role, reduce the release of inflammatory factors, and alleviate the damage to the back skin of AD mice.

[0114] Experiment 2: Experimental study on the effect of the composition of Example 1 on promoting wound healing in scalded mice

[0115] Scalds are injuries caused by hot liquids, solids, or steam. Scalds are categorized into three degrees: first-degree burns damage only the surface of the skin, causing mild redness and swelling without blisters; second-degree burns damage the dermis, with blisters of varying sizes; and third-degree burns injure the subcutaneous layer, affecting the fat, muscle layer, and bone. Currently, small, mild burns should be immediately rinsed with room-temperature water for several minutes, followed by disinfection and bandaging. Small, deep burns should be disinfected and bandaged, followed by skin grafting to promote wound recovery, depending on the patient's condition. Larger burns require fluid replacement within 24 hours after medical treatment to maintain fluid balance, keep the wound clean, and regularly change dressings and bandages. Depending on the severity of the burn and recovery, one or more skin grafts may be performed. Example 2 evaluated the improvement of an OSHP formulation combination on scalded mice by establishing a scald mouse model, providing data support for the application of an OSHP formulation combination with a buffered pH value in functional products.

[0116] 1. Reagent Preparation

[0117] Preparation of 2% sodium alginate: 1 g of sodium alginate was dissolved in 50 ml of water and placed on a magnetic stirrer to dissolve.

[0118] OSHP prescription combination gel: effective concentration 100mg / ml, dissolved in 2% sodium alginate.

[0119] 2. Animal burn modeling, grouping and medication

[0120] The experimental group was mainly divided into 3 groups, model group, empty load group (gel was applied only on the affected area), and OSHP prescription combination group. Use an electric shaver to shave 2.5cm×2.5cm of the mouse's back skin, remove the mice with problems on the back skin to avoid interference, and use depilatory cream to remove hair twice on the shaved area to ensure that the mouse's back skin can be completely exposed. After using the depilatory cream, wash it with water to prevent the depilatory cream from corroding the mouse skin and affecting the modeling effect. On the first day of the experiment, all three groups of mice were modeled under anesthesia using a scalding instrument. Drug treatment began on the day of modeling, and the drug concentration of the OSHP prescription combination group was 100mg / ml. All drugs were administered externally to ensure that the drug efficacy was fully exerted. The drug was administered for a total of 26 days, and experimental samples were collected on the 26th day.

[0121] 3. Healing of back wounds of scalded mice

[0122] In this experiment, the wound surfaces of the mice were photographed every other day to compare the surface healing effects. The changes in the wound healing status of the mice were used to determine the efficacy of a pH-buffered oyster shell hydrogen powder combination.

[0123] like Figure 7 As shown, the OSHP combination, at a concentration of 100 mg / ml, significantly improved the healing of burn wounds in mice. Wounds in the model and empty-vessel groups completely healed by day 20, while those in the OSHP combination group healed by day 14, a week shorter than in the other groups. This may be due to the hydrogen released by OSHP in the mouse wounds, which reduces the release of inflammatory factors, accelerates burn wound healing, and repairs the mouse skin, promoting skin growth and healing. This demonstrates that an OSHP combination with a buffered pH can rapidly promote wound healing.

[0124] 4. Changes in mouse weight

[0125] In this experiment, mice were weighed daily. Changes in their body weight were used to assess the efficacy of a pH-buffered OSHP formulation on wound healing in scalded mice.

[0126] Mouse weight is an important indicator for evaluating mouse growth and development, as well as an evaluation indicator for mouse quality of life and disease recovery. Figure 8 As shown in the figure, the weight growth curve of scalded mice showed that the weight growth trend of mice in the OSHP prescription combination group was better than that in other groups. This may be related to the faster wound healing speed of mice in the OSHP prescription combination group, better disease recovery of mice, and improved quality of life due to reduced pain caused by disease recovery.

[0127] The oyster shell hydrogen powder composition with pH buffering function of the present invention has significant anti-inflammatory and wound healing effects, and can provide the body with a stable and long-term supply of hydrogen. Oral administration of the composition can effectively increase the H2 content in the human body, thereby exerting anti-inflammatory effects and promoting wound growth and healing. The composition of the present invention also has the advantages of high safety and minimal side effects, making it suitable for long-term use by a wide range of people.

[0128] The present invention utilizes solid-state hydrogenation technology to produce a hydrogen-rich OSHP, which slowly and continuously releases hydrogen upon contact with water. This allows for the slow and continuous release of hydrogen on the body surface after external application, promoting rapid wound healing.

[0129] An H2-rich oyster shell powder produced by solid-state hydrogenation technology has the characteristic of slowly and continuously releasing H2 when it comes into contact with water. Oral administration through the gastrointestinal tract can slowly and continuously release H2 with certain health benefits, thereby achieving significant anti-inflammatory, immune regulation, and metabolic regulation effects.

[0130] The present invention uses OSHP, citric acid, hydroxyethyl methylcellulose (HEMC), and starch as ingredients in an OSHP formulation with a buffered pH value. Taking advantage of OSHP's ability to release hydrogen over a long period of time, citric acid (CA) is added for acid-base buffering, thereby adjusting the pH value of the OSHP formulation to 7, which is effective in combating inflammation, regulating immunity and metabolism, and promoting rapid wound healing. Hydroxyethyl methylcellulose (HEMC) is further added to improve drug stability and enhance sustained-release effects, thereby enhancing drug safety.

Claims

1. An oyster shell hydrogen powder composition having a pH buffering function, characterized in that: The composition is as follows by mass percentage: 40% to 50% oyster shell hydrogen powder, 20% to 30% starch, 10% to 15% citric acid, and 15% to 20% hydroxyethyl methyl cellulose, and the sum of the percentages of the above components is 100%; The citric acid acts as an acidity regulator to adjust the pH value of the composition to a stable neutral state; The hydroxyethyl methylcellulose is used as a sustained-release preparation to improve the stability of the composition and enhance the safety of the composition.

2. The oyster shell hydrogen powder composition with pH buffering function according to claim 1, characterized in that: The composition is as follows by mass percentage: 45% oyster shell hydrogen powder, 24% starch, 13% citric acid, and 18% hydroxyethyl methylcellulose.

3. The oyster shell hydrogen powder composition with pH buffering function according to claim 1 or 2, characterized in that: The oyster shell hydrogen powder composition is packaged in capsule shells made of edible and easily digestible gelatin, and is made into capsules with a specification of 0.45 g of content per capsule.

4. The method for preparing the oyster shell hydrogen powder composition having a pH buffering function according to claim 3, characterized in that: Prepare as follows: Step 1: Pass oyster shell hydrogen powder, starch, citric acid, and hydroxyethyl methylcellulose through a 100-mesh sieve respectively for later use; Step 2: According to the formula, accurately weigh the oyster shell hydrogen powder, starch, citric acid, and hydroxyethyl methylcellulose and mix them together; Step 3, filling the total mixed material into hollow capsules; Step 4: Pack the filled capsules.

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